BETA-GALACTOSIDASE ACTIVITY IN SINGLE DIFFERENTIATING BACTERIAL-CELLS
BETA-GALACTOSIDASE ACTIVITY IN SINGLE DIFFERENTIATING BACTERIAL-CELLS
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DOI：
10.1073/pnas.90.17.8194
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发表时间：
1993-09-01
影响因子：
11.1
通讯作者：
KAISER, D
中科院分区：
文献类型：
--
作者：
RUSSOMARIE, F;ROEDERER, M;KAISER, D
对含有与lacZ的转录融合的黄色粘球菌菌株进行了分析，并通过其β-半乳糖苷酶表达水平的差异进行了分级。 将β-半乳糖苷酶的荧光底物荧光素二-β-吡喃半乳糖苷引入M.在培养基的渗透压迅速降低，随后恢复到等渗浓度的过程中，水解产物黄绿素保留在细胞内，并保持其活力。荧光随时间线性增加，并与β-半乳糖苷酶活性成正比。 β-半乳糖苷酶的表达在大多数融合株，虽然开始在不同阶段的生长或发展，是单峰分布在细胞之间。然而，融合菌株Tn 5 lac OMEGA 4473显示在9小时发育时是异质的。有可能将高水平表达β-半乳糖苷酶的细胞与其他仍存活的无表达细胞物理分离。这里描述的方法可以适用于研究植物和动物的分化，也可以使用基因表达的酶探针的转录融合和荧光底物。
Myxoccus xanthus strains containing transcriptional fusions to lacZ were analyzed and fractionated by differences in their levels of beta-galactosidase expression. The fluorogenic substrate for beta-galactosidase, fluorescein di-beta-galactopyranoside, was introduced into M. xanthus cells during a rapid decrease in osmolarity of the medium followed by a return to isoosmolarity. Fluorescein, the product of hydrolysis, was retained within the cells and their viability was preserved. Fluorescence increased linearly with time and was proportional to beta-galactosidase activity. Beta-galactosidase expression in most fusion strains, though beginning at different phases of growth or development, was distributed unimodally amongst cells. However, fusion strain Tn5 lac OMEGA4473 was shown to be heterogeneous at 9 hr of development. It was possible to separate physically cells that expressed beta-galactosidase at a high level from other, still viable, cells with no expression. The approach described here could be adapted to study differentiation in plants and animals as well, where transcriptional fusions and fluorogenic substrates for enzyme probes of gene expression also can be used.